LOCUS       sequence-182037-        3241 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of NTerm 6xHis tagged mRFP mutant K15TAG for ncAA 
            incoporation using the p70a promoter and UTR1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3241)
  AUTHORS   Schlesinger O, Chemla Y, Heltberg M, Ozer E, Marshall R, Noireaux V,
            Jensen MH, Alfonta L
  TITLE     Tuning of Recombinant Protein Expression in Escherichia coli by 
            Manipulating Transcription, Translation Initiation Rates, and 
            Incorporation of Noncanonical Amino Acids.
  JOURNAL   ACS Synth Biol. 2017 Mar 9. doi: 10.1021/acssynbio.7b00019.
  PUBMED    28230975
REFERENCE   2  (bases 1 to 3241)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "ACS Synth 
            Biol. 2017 Mar 9. doi: 10.1021/acssynbio.7b00019."
COMMENT     Sequence Label: pBEST-p70a-UTR1-Nterm 6xHis mRFP K15TAG-T500
FEATURES             Location/Qualifiers
     source          1..3241
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     444..463
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(682..786)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     854..872
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     RBS             1210..1232
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1251..1268
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             1278..1955
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRF*VRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     primer_bind     complement(2132..2149)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2303..2891)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2383..2402)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3062..3241,1..612,613..681))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
ORIGIN
        1 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
       61 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      121 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      181 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      241 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      301 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      361 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      421 ctcttgcccg gcgtcaacac gggataatac cgcgccacat agcagaactt taaaagtgct
      481 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      541 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      601 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      661 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      721 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      781 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
      841 attaacctat aaaaataggc gtatcacgag gccctttcgt cttcaagaat tctggcgaat
      901 cctctgacca gccagaaaac gacctttctg tggtgaaacc ggatgctgca attcagagcg
      961 ccagcaagtg ggggacagca gaagacctga ccgccgcaga gtggatgttt gacatggtga
     1021 agactatcgc accatcagcc agaaaaccga attttgctgg gtgggctaac gatatccgcc
     1081 tgatgcgtga acgtgacgga cgtaaccacc gcgacatgtg tgtgctgttc cgctgggcat
     1141 gctgagctaa caccgtgcgt gttgacaatt ttacctctgg cggtgataat ggttgcagct
     1201 agcaataatt ttgtttaact ttaagaagga gatataccat gggcagcagc catcatcatc
     1261 atcatcacag ctctagaatg gcttcctccg aagacgttat caaagagttc atgcgtttct
     1321 aggttcgtat ggaaggttcc gttaacggtc acgagttcga aatcgaaggt gaaggtgaag
     1381 gtcgtccgta cgaaggtacc cagaccgcta aactgaaagt taccaaaggt ggtccgctgc
     1441 cgttcgcttg ggacatcctg tccccgcagt tccagtacgg ttccaaagct tacgttaaac
     1501 acccggctga catcccggac tacctgaaac tgtccttccc ggaaggtttc aaatgggaac
     1561 gtgttatgaa cttcgaagac ggtggtgttg ttaccgttac ccaggactcc tccctgcaag
     1621 acggtgagtt catctacaaa gttaaactgc gtggtaccaa cttcccgtcc gacggtccgg
     1681 ttatgcagaa aaaaaccatg ggttgggaag cttccaccga acgtatgtac ccggaagacg
     1741 gtgctctgaa aggtgaaatc aaaatgcgtc tgaaactgaa agacggtggt cactacgacg
     1801 ctgaagttaa aaccacctac atggctaaaa aaccggttca gctgccgggt gcttacaaaa
     1861 ccgacatcaa actggacatc acctcccaca acgaagacta caccatcgtt gaacagtacg
     1921 aacgtgctga aggtcgtcac tccaccggtg cttaactcga gcaaagcccg ccgaaaggcg
     1981 ggcttttctg tgtcgaccga tgcccttgag agccttcaac ccagtcagct ccttccggtg
     2041 ggcgcggggc atgactatcg tcgccgcact tatgactgtc ttctttatca tgcaactcgt
     2101 aggacaggtg ccggcagcgc tcttccgctt cctcgctcac tgactcgctg cgctcggtcg
     2161 ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta tccacagaat
     2221 caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc aggaaccgta
     2281 aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa
     2341 atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc
     2401 cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt
     2461 ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca
     2521 gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg
     2581 accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat
     2641 cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta
     2701 cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct
     2761 gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac
     2821 aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa
     2881 aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa
     2941 actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt
     3001 taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca
     3061 gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca
     3121 tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc
     3181 ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa
     3241 a
//
